PROJECT SUMMARY/ABSTRACT The overarching goal of my laboratory is to determine how natural genetic variation influences chromatin biology, and, ultimately, phenotypic diversity. Most disease-associated variants identified in human studies occur in regulatory elements rather than in the coding region of genes, highlighting the importance of understanding the role of regulatory variation in normal health and disease. Gene transcription is controlled by the interplay between cis-acting regulatory elements and the trans-acting factors that bind them. Variation within a regulatory element can influence function locally through disruption of a DNA-binding site targeted by a trans-acting factor at that site, whereas variation in sequence or expression of a trans-acting factor can result in functional variation distally at many regulatory elements. Regulatory elements are identified by epigenetic features catalyzed by a variety of chromatin writers. While we have substantial information on the enzymes that write and erase epigenetic modifications, less is known about the trans-acting mechanisms that regulate the locations and levels of these modifications, remaining a defining challenge within the field. To move the field forward, we have developed a novel cellular systems genetics approach that integrates quantitative experimental techniques and computational methods in statistical genetics to create a comprehensive understanding of regulatory variation and uncover broadly applicable mechanisms that control the epigenetic landscape. We will utilize the natural genetic variation intrinsic in a unique panel of mouse embryonic stem cells that will enable us to harness genetic diversity to identify key chromatin regulators, define their molecular functionality across multiple phenotypes, and delineate changes in regulatory variation over developmental time. This mammalian genetic reference panel will greatly improve statistical power to efficiently map functional loci directly relatable to humans, whereas a cellular platform will enable the identification of their associated genes and mechanisms of control. Using this system we have discovered multiple genomic locations that distally control hundreds of regulatory elements and alter gene expression. Our goals in the next five years are to answer the following questions: What are the molecules and mechanisms underlying trans regulation of the chromatin landscape? How does genetic variation influence chromatin structure and function? How does the early establishment of the chromatin landscape impact development? Success of this project will delineate the interplay between regulatory elements, the systems that control them, and the epigenetic landscape that defines them. These efforts are expected to have broad implications in both our basic understanding of chromatin and epigenetic regulation during early development as well as on our ability to directly tailor in vitro cellular differentiation to genetic background, ultimately impacting developmental biology, regenerative biology and personalized medicine.